Molecular redundant sequencing

Abstract

Methods, systems and compositions where a target nucleic acid includes a registration sequence disposed therein for identification of the number or relative position of determined sequence from the template sequence. Particularly preferred aspects include a registration sequence in a circular template nucleic acid sequence which is, in turn, used in sequence by incorporation processes that rely upon template dependent, polymerase mediated primer extension in the identification of the sequence of the template.

What is claimed is: 
     
       1. A method of sequencing a target sequence, comprising:
 providing a linear nucleic acid template comprising multiple copies of the target sequence, wherein a registration sequence is identically located within each of the multiple copies of the target sequence; 
 sequencing the nucleic acid template until at least three of the multiple copies of the target sequence have been sequenced, thereby providing at least three sequence reads for the target sequence; 
 aligning the sequence reads to one another based at least in part on the location of the registration sequence in each of the sequence reads, thereby providing aligned sequence reads; and 
 determining a consensus sequence from the aligned sequence reads by calling bases of the target sequence based upon at least a 60% consensus among the sequence reads. 
 
     
     
       2. The method of  claim 1 , wherein the providing step comprises performing rolling circle replication of a circular nucleic acid comprising the target sequence and the registration sequence. 
     
     
       3. The method of  claim 1 , wherein the sequencing process comprises identifying bases upon incorporation in a template directed, polymerase mediated primer extension reaction. 
     
     
       4. The method of  claim 3 , wherein the incorporation occurs in the presence of four different nucleotide analogs. 
     
     
       5. The method of  claim 4 , wherein the four different nucleotide analogs are differentially labeled. 
     
     
       6. The method of  claim 1 , wherein the sequencing process comprises a Sanger sequencing process. 
     
     
       7. The method of  claim 1 , wherein the sequencing process comprises a real-time sequencing process. 
     
     
       8. The method of  claim 1 , wherein said sequencing is nondestructive to the nucleic acid template. 
     
     
       9. The method of  claim 1 , wherein said determining is based upon at least a 70% consensus among the sequence reads. 
     
     
       10. The method of  claim 1 , wherein said determining is based upon at least a 80% consensus among the sequence reads. 
     
     
       11. The method of  claim 1 , wherein said sequencing provides at least four sequence reads for the target sequence. 
     
     
       12. The method of  claim 1 , wherein said sequencing provides at least five sequence reads for the target sequence. 
     
     
       13. The method of  claim 1 , wherein said sequencing provides at least ten sequence reads for the target sequence. 
     
     
       14. The method of  claim 1 , wherein the target sequence comprises at least 100 bases. 
     
     
       15. The method of  claim 1 , wherein the linear nucleic acid template comprises at least 1000 bases. 
     
     
       16. The method of  claim 1 , wherein the aligning comprises positioning the registration sequence in each of the sequence reads in the same sequence location.